New mechanism of chromatin compartmentalization by BRD2
Authors: Yubao Cheng, Siyuan Wang
Journal: Trends in Genetics (2022)

The molecular mechanism underlying 3D genome compartmentalization remains a mystery. Xie et al. found that the bromodomain and extraterminal (BET) family scaffold protein BRD2 promotes the compartmentalization of the accessible chromatin domains after cohesin loss. An antagonistic interplay between loop extrusion and compartmentalization modulates chromatin folding. The molecular mechanism underlying 3D genome compartmentalization remains a mystery. Xie et al. found that the bromodomain and extraterminal (BET) family scaffold protein BRD2 promotes the compartmentalization of the accessible chromatin domains after cohesin loss. An antagonistic interplay between loop extrusion and compartmentalization modulates chromatin folding. Interphase chromatin folds hierarchically in mammalian cells. Topologically associating domains (TADs, also known as contact domains) and A–B compartments are two major architectures of the chromatin-folding hierarchy [1.Dixon J.R. et al.Topological domains in mammalian genomes identified by analysis of chromatin interactions.Nature. 2012; 485: 376-380Crossref PubMed Scopus (4097) Google Scholar, 2.Nora E.P. et al.Spatial partitioning of the regulatory landscape of the X-inactivation centre.Nature. 2012; 485: 381-385Crossref PubMed Scopus (1873) Google Scholar, 3.Lieberman-Aiden E. et al.Comprehensive mapping of long-range interactions reveals folding principles of the human genome.Science. 2009; 326: 289-293Crossref PubMed Scopus (5055) Google Scholar]. TADs are local chromatin domains that are tens to hundreds of kilobases in size, and have more intradomain than interdomain chromatin contacts [1.Dixon J.R. et al.Topological domains in mammalian genomes identified by analysis of chromatin interactions.Nature. 2012; 485: 376-380Crossref PubMed Scopus (4097) Google Scholar,2.Nora E.P. et al.Spatial partitioning of the regulatory landscape of the X-inactivation centre.Nature. 2012; 485: 381-385Crossref PubMed Scopus (1873) Google Scholar]. At the chromosome scale, these self-interacting domains are further sorted into A and B compartments, enriched with active and inactive chromatin, respectively [3.Lieberman-Aiden E. et al.Comprehensive mapping of long-range interactions reveals folding principles of the human genome.Science. 2009; 326: 289-293Crossref PubMed Scopus (5055) Google Scholar]. The chromatin domains in the same compartment interact over a long distance and are spatially grouped together in individual copies of chromosomes [4.Wang S.Y. et al.Spatial organization of chromatin domains and compartments in single chromosomes.Science. 2016; 353: 598-602Crossref PubMed Scopus (341) Google Scholar]. The mechanism of TAD formation has been extensively studied under a loop extrusion model [5.Davidson I.F. Peters J.M. Genome folding through loop extrusion by SMC complexes.Nat. Rev. Mol. Cell Biol. 2021; 22: 445-464Crossref PubMed Scopus (117) Google Scholar]: cohesin complexes extrude DNA loops that correspond to TADs, and stall at TAD boundaries where the complexes are blocked by CTCF proteins bound to the boundary DNA locus in an effective orientation. After rapid degradation of cohesin, the A and B compartments persist and are even strengthened, suggesting a cohesin-independent and counteractive role of A–B compartmentalization [6.Rao S.S.P. et al.Cohesin loss eliminates all loop domains.Cell. 2017; 171: 305-320Abstract Full Text Full Text PDF PubMed Scopus (903) Google Scholar]. Phase-separating interaction between heterochromatic regions was suggested to contribute to compartment B formation [7.Falk M. et al.Heterochromatin drives compartmentalization of inverted and conventional nuclei.Nature. 2019; 570: 395-399Crossref PubMed Scopus (269) Google Scholar], while mechanistic details require further investigation [8.Erdel F. et al.Mouse heterochromatin adopts digital compaction states without showing hallmarks of HP1-driven liquid-liquid phase separation.Mol. Cell. 2020; 78: 236-249Abstract Full Text Full Text PDF PubMed Scopus (128) Google Scholar,9.McSwiggen D.T. et al.Evaluating phase separation in live cells: diagnosis, caveats, and functional consequences.Genes Dev. 2019; 33: 1619-1634Crossref PubMed Scopus (280) Google Scholar]. The molecular mechanism underlying the formation of the compartment A has remained largely unclear. A new study by Xie et al. now sheds light on this question. Using a super-resolution imaging method, they first visualized the spatial organization of accessible chromatin domains (ACDs) upon cohesin loss by rapid depletion of cohesin subunit RAD21 [10.Xie L. et al.BRD2 compartmentalizes the accessible genome.Nat. Genet. 2022; 54: 481-491Crossref PubMed Scopus (7) Google Scholar]. The ACDs are expected to be enriched in compartment A. They found that cohesin loss triggered spatial mixing of ACDs, in agreement with the strengthened compartment. To explore the underlying mechanism, they further perturbed the cells with several small-molecule inhibitors. Interestingly, JQ1, an inhibitor of the BET protein family (including BRD2, BRD3, BRD4, and BRDT), reduced the spatial mixing and induced decompaction of ACDs after cohesin loss. BET family proteins were known to be transcriptional coregulators that bind to active chromatin. To further dissect the role of BET family proteins in genome compartmentalization, Xie et al. performed combined depletion of RAD21 with BRD2/3/4. Surprisingly, only BRD2 depletion reversed ACD mixing and induced the decompaction of ACDs. BRD2 depletion and overexpression also reduced and enhanced chromatin contacts in compartment A, respectively [10.Xie L. et al.BRD2 compartmentalizes the accessible genome.Nat. Genet. 2022; 54: 481-491Crossref PubMed Scopus (7) Google Scholar]. To further understand the role of BRD2 in active chromatin compartmentalization, Xie et al. examined the chromatin-binding dynamics of BET family proteins by single-molecule tracking [10.Xie L. et al.BRD2 compartmentalizes the accessible genome.Nat. Genet. 2022; 54: 481-491Crossref PubMed Scopus (7) Google Scholar]. Cohesin loss led to a higher chromatin-bound fraction and decreased the diffusion coefficient of BRD2, but not of BRD3 or BRD4. Chromatin immunoprecipitation followed by sequencing (ChIP-seq) and biochemical fractionation experiments further confirmed the stronger chromatin binding of BRD2 after cohesin loss. These results support an antagonistic relationship between BRD2 and cohesin [10.Xie L. et al.BRD2 compartmentalizes the accessible genome.Nat. Genet. 2022; 54: 481-491Crossref PubMed Scopus (7) Google Scholar]. The unexpected role of BRD2 in chromatin organization led Xie et al. to further investigate the interplay between BET family proteins [10.Xie L. et al.BRD2 compartmentalizes the accessible genome.Nat. Genet. 2022; 54: 481-491Crossref PubMed Scopus (7) Google Scholar]. They performed depletion of BRD2/3/4 individually followed by ChIP-seq. BRD2 and BRD4 appeared to antagonize each other, because the depletion of either resulted in enhanced chromatin binding of the other. From cell imaging, the increased fluorescence intensity of labeled BRD2 puncta after BRD4 depletion further indicated their competition in chromatin binding. Dual depletion of BRD4 and cohesin led to more contacts and stronger clustering of activate chromatin compared with cohesin depletion alone, which could be reversed by further BRD2 depletion. BRD4, but not BRD2, depletion significantly reduced the chromatin binding of transcriptional co-activating protein P300, coupled with differential expression of thousands of genes. These results demonstrated distant functions of BRD2 and BRD4, although they have similar protein domains and binding targets and both form puncta in the cell nucleus. The observations suggest that the BRD4 puncta are transcription hotspots, while the BRD2 puncta are hubs of chromatin interaction in compartment A (Figure 1A ). Finally, by simulating loop extrusion in combination with BRD2-induced interaction of activate chromatin, Xie et al. successfully recapitulated several key experimental observations [10.Xie L. et al.BRD2 compartmentalizes the accessible genome.Nat. Genet. 2022; 54: 481-491Crossref PubMed Scopus (7) Google Scholar]. The work by Xie et al. reveals a novel molecular mechanism that directly contributes to the formation of compartment A in 3D genome organization. BRD4 and several other transcription-related chromatin-binding proteins were previously hypothesized to drive compartment A formation due to their phase-separation properties. Importantly, this new work shows that the chromatin-binding and phase-separation properties of a protein may not guarantee a positive role of the protein in chromatin compartmentalization. The work comprehensively demonstrated how the competitive interplay between BRD2, cohesin, and BRD4 balance higher-order chromatin organization. Besides the contrasting roles of BRD2 and BRD4, one seeming similarity between them is that the depletion of either leads to the conversion of some compartment B regions to compartment A, except that the effect by BRD2 depletion depends on cohesin, while the effect of BRD4 depletion was shown on a background of cohesin loss [10.Xie L. et al.BRD2 compartmentalizes the accessible genome.Nat. Genet. 2022; 54: 481-491Crossref PubMed Scopus (7) Google Scholar]. Xie et al. explained the BRD2 effect on the compartment conversion by a model in which the BRD2–CTCF interaction safeguards the TAD boundary between compartment A and B regions (Figure 1B). A hidden prerequisite of the model is that, upon BRD2 loss, the loop extrusion events from compartment A should overwhelm loop extrusion from compartment B and, in turn, reel in the ex-compartment-B region across the weakened boundary into compartment A. This stronger or more frequent loop extrusion in compartment A may be caused by a BRD4 interaction with cohesin loader protein NIPBL [10.Xie L. et al.BRD2 compartmentalizes the accessible genome.Nat. Genet. 2022; 54: 481-491Crossref PubMed Scopus (7) Google Scholar]. This model predicts that the converted regions should be adjacent to pre-existing compartment A regions. The cohesin-independent effect of BRD4 depletion on the compartment conversion can be explained by: (i) increased binding of BRD2 to cryptic compartment A regions previously embedded in compartment B, which converts these cryptic A regions to definitive A regions (Figure 1C); and/or (ii) enhanced clustering of pre-existing compartment A regions by BRD2, which sequesters adjacent compartment B regions into compartment A (Figure 1D). While the two models are not mutually exclusive, the second predicts the converted regions to be adjacent to pre-existing compartment A regions. Further studies are needed to fully reveal the molecular interplay among BRD2, BRD4, loop extrusion machinery, and chromatin. It remains to be discovered what other factors contribute to A–B compartmentalization and whether they drive compartmentalization through a phase-separation mechanism. Y.C. declares no conflicts of interest. S.W. is a consultant and shareholder of Translura, Inc.